The Krebs tricarboxylic acid cycle (the “TCA cycle”) and oxidative phosphorylation are central to metabolic energy production. The TCA cycle occurs in the mitochondria of cells and, in most cells, produces the majority of adenosine triphosphate (>90%). In normal cells, the main energy source for the TCA cycle is pyruvate generated from glycolysis of glucose.
Many disease states have perturbed TCA cycles. In cancer, succinate dehydrogenase and fumarate hydratase oncogenes impair the TCA cycle. The TCA cycle can have different entry points. A broad range of energy substrates can be used in the TCA cycle (e.g., citrate and glutamate/glutamine), especially in cancer. Evidence exists that the TCA cycle is altered in many neurodegenerative diseases (e.g., Alzheimer's, Parkinson's, Huntington's, and amyotrophic lateral sclerosis) as well. In addition, adenosine triphosphate (“ATP”) is the main energy source for the heart for contraction, maintenance of active ion gradients, and other vital functions. Most of the ATP production of the heart occurs as a result of, and is controlled by, the TCA cycle, and the TCA cycle is altered in many disease states of the heart.
Many of the metabolic differences between disease and normal tissue can and could be examined through the use of metabolic imaging agents. Presently, metabolic imaging is performed with positron emission tomography (“PET”) measurements of the uptake of 2-[18F]fluoro-2-deoxy-D-glucose (“FDG-glucose”) or magnetic resonance spectroscopy (“MRS”). However, PET imaging with FDG-glucose only measures the level of uptake of glucose and phosphorylation, and reveals nothing about the subsequent metabolism of glucose. In MRS, only the steady state of a tissue/organ's metabolic profile can be determined, and, under normal circumstances, MRS's low signal to noise ratio requires lengthy exam times.
More recently, hyperpolarization of molecules has opened the way to real-time metabolic imaging in vivo, i.e., in living human or non-human animal beings. Hyperpolarization allows for over 10,000 fold sensitivity enhancement using conventional magnetic resonance imaging (“MRI”) and MRS. Upon enhancing the nuclear polarization of nuclear magnetic resonance (“NMR”) active nuclei (e.g., 13C, 15N, 31P), the population difference between excited and ground nuclear spin states of the nuclei is increased and the magnetic resonance signal intensity is amplified. The polarization (signal enhancement) may be retained on the metabolites of the hyperpolarized molecule. In addition, unlike PET, the hyperpolarization process is non-radioactive.
The most widely used methods for hyperpolarization are dynamic nuclear polarization (“DNP”) and parahydrogen induced polarization (“PHIP”). Several compounds have been hyperpolarized and studied using hyperpolarized metabolic imaging. For example, 1-13C pyruvate, 1,4-13C fumarate, 13C succinate, 13C 2-hydroxy ethyl propionate, and 2,2,3,3-tetrafluoropropyl-1-13C priopionate-d2,2,3,3 (“TFPP”) have been studied in in vivo applications. However, all of these compounds have physiological barriers to being used in clinical practice. For example, 1-13C pyruvate can be used to follow the metabolism of pyruvate to alanine, lactate, and bicarbonate, but reveals nothing about TCA cycle metabolism. For 13C succinate, the polarization transfer must be performed under acidic (pH≦3) or alkaline (pH≧9) conditions for optimum hyperpolarization. In addition, 13C succinate is only poorly transported across many biological membranes and, in particular, barely crosses the mitochondrial membrane to gain access to TCA cycle enzymes involved in metabolism. 13C 2-hydroxy ethyl propionate is toxic and is not metabolized. TFPP is not very water soluble and has to be injected in a 20% ethanol aqueous solution.
Hyperpolarized compounds are needed that: (1) are metabolized by the TCA cycle; and (2) are capable of use as diagnostic in vivo imaging agents.